Unlike the hearts of teleosts or zebrafish,1
the mammalian heart undergoes a fibrotic response with minimal regeneration.2
The inability of the adult heart to completely repair itself spurs the development of strategies to transplant endogenously or exogenously-derived cardiac cells into patients after ischemic injury3–8
. These transplantation strategies have led to measurable successes in functional recovery or remuscularization. However, significant challenges remain regarding the efficiency, feasibility and efficacy of this approach6–9
. Thus, strategies aimed at directly converting cardiac scar fibroblasts into cardiomyocytes are appealing as they circumvent problems associated with cell purity and survival after transplantation.
To this end, Ieda and colleagues reported that overexpression of Gata4, Mef2c and Tbx5 (GMT) could reprogram murine cardiac fibroblasts (CFs) and tail tip fibroblasts (TTFs) into cardiomyocytes in vitro.10
Furthermore, infected fibroblasts could survive and reprogram after transplantation into a murine heart. GMT-induced fibroblasts demonstrated gene expression profiles similar to mature cardiomyocytes and beat spontaneously in vitro
. Conversion to cardiomyocyte-like epigenetic states was reported through the de-repression of histone markings at promoters of sarcomeric genes. These results implicated therapies that can directly remuscularize the heart without the need for cell transplantation, provided that the efficiency of reprogramming is sufficiently robust.
Here, we evaluated the efficiency of this direct cardiac reprogramming strategy using the GMT expression viruses reported in Ieda et al.10
and validated myocardial lineage reporters (αMHC-Cre, Nkx2.5-Cre, cTnT-Cre). We found a lack of αMHC or Nkx2.5 reporter activation in CFs and TTFs despite significant overexpression of GMT factors. However, with cTnT reporter we observed a ~35% labeling of fibroblasts that is confirmed by a ~250-fold increase in cTnT expression. However, the expression of other cardiac genes was minimally elevated. With GMT infection, we found 22% of infected fibroblasts exhibited a voltage-dependent calcium current without a spontaneous action potential, suggesting incomplete electrophysiological reprogramming. Furthermore, GMT-infected fibroblasts exhibited poor survival and minimal cardiac gene expression following transplantation into an injured murine heart in vivo
. Together, our data suggest that direct cardiomyocyte reprogramming by GMT factors is inefficient and a greater understanding of transcription factor-mediated epigenetic change will be need to translate this promising approach into therapy.